Defining in vivo transcriptional responses to auxin

Abstract The plant hormone auxin regulates many aspects of the plant life cycle through a series of transcriptional responses mediated by Auxin Response Factors (ARFs). However, thus far, it has not been possible to identify in vivo DNA binding sites for most of ARFs leading to a major gap in our understanding of these master regulatory transcription factors. Here we identify the DNA binding profiles for several ARFs as well as Aux/IAA proteins (IAAs), repressors of the response to auxin that heterodimerize with ARFs, using ChIP-seq. As expected, the common target genes of all tested ARFs are highly enriched in known auxin-responsive genes, each containing the motif K(T/G)GTCBB(T/G/C) with a core GTC and a non-A flanking sequence. Unexpectedly, genome-wide ARF binding sites are strongly associated with Aux/IAA ChIP-seq peaks, providing evidence that the Aux/IAA-ARF complex may control auxin transcriptional responses directly on the promoters of hormone-responsive genes. Additionally, we find that ABA response genes are also primary targets of an ARF10-IAA19 module and that a putative repressor ARF(ARF10) binds the same set of targets as activator ARFs. Finally, although ARFs have previously been reported to have strict spacing between their motifs when assayed in vitro, we observe that conservation of the spacing of ARF binding motifs is less rigid when assayed in planta.